Expression of BAG1 is associated with prognosis in kidney renal clear cell carcinoma based on bioinformatics.

BACKGROUND: BCL2 associated Athano-Gene 1 (BAG1) has been described to be involved in the development and progression of cancer. But the role of BAG1 in kidney renal clear cell carcinoma (KIRC) has remained largely unknown. METHODS: We performed bioinformatic analysis of data from TCGA and GEO dataset. The role of BAG1 in KIRC was explored by Logistic and Cox regression model. The molecular mechanisms of BAG1 was revealed by GSEA. RESULTS: The current study found that the KIRC tumor samples have a low level of BAG1 mRNA expression compared to the matched normal tissues based on TCGA data and GEO databases. Low expression of BAG1 in KIRC was significantly associated with Sex, clinical pathological stage, tumor-node-metastasis (TNM) stage, hemoglobin levels, cancer status and history of neoadjuvant treatment. Kaplan-Meier survival analysis indicated that KIRC patients with BAG1 high expression have a longer survival time than those with BAG1 low expression (p < 0.000). Cox regression analysis showed that BAG1 remained independently associated with overall survival, with a hazard ratio (HR) of 1.75(CI:1.05-2.90; p = 0.029). GSEA indicated that the signaling pathways including fatty acid metabolism and oxidative phosphorylation were differentially enriched in high BAG1 expression phenotype. CONCLUSIONS: These findings suggested that BAG1 expression may act as a potential favorable prognostic marker and challenging therapeutic target.

SOCS1 Regulates the Immunomodulatory Roles of MSCs on B Cells.

BACKGROUND AND OBJECTIVES: The effective use of MSCs for the treatment of some B cell-mediated immune diseases is quite limited. The main reason is that the immunomodulatory effects of mesenchymal stem cells (MSCs) on B cells are unclear, and their underlying mechanisms have not been fully explored. METHODS AND RESULTS: By co-culturing B cells with MSCs without (MSC/CTLsh) or with suppressor of cytokine signaling 1 (SOCS1) knockdown (MSC/SOCS1sh), we found that MSCs inhibited B cell proliferation, activation and terminal differentiation. Remarkably, the highest inhibition of B cell proliferation was observed in MSC/SOCS1sh co-culture. Besides, MSC/SOCS1sh reversed the inhibitory effect of MSCs in the last stage of B cell differentiation. However, MSC/SOCS1sh had no effect on inhibiting B cell activation by MSCs. We also showed that IgA+ B cell production was significantly higher in MSC/SOCS1sh than in MSC/CTLsh, although no difference was observed when both MSCs co-cultures were compared to isolated B cells. In addition, MSCs increased PGE2 production after TNF-α/IFN-γ stimulation, with the highest increase observed in MSC/SOCS1sh co-culture. CONCLUSIONS: Our results highlighted the role of SOCS1 as an important new mediator in the regulation of B cell function by MSCs. Therefore, these data may help to develop new treatments for B cell-mediated immune diseases.

In vivo Therapeutic Effects and Mechanisms of Hydroxyasiaticoside Combined With Praziquantel in the Treatment of Schistosomiasis Induced Hepatic Fibrosis.

Schistosomiasis has been a fatal obstinate disease that threatens global human health, resulting in the granulomatous inflammation and liver fibrosis. Objective:The aim of this study was to evaluate the therapeutic effects and mechanisms of hydroxyasiaticoside combined with praziquantel in the treatment of schistosomiasis-induced liver fibrosis. Methods:Mice were randomly distributed into four experimental groups: normal control group, model group, praziquantel group, praziquantel + hydroxyasiaticoside group. Except for the normal control group, they were infected with Schistosomia cercariae through the abdominal skin to induce liver fibrosis. In the intervention group, mice were administered with the respective drugs by gavage after 8 weeks of infection. At the end of the treatment, mice were sacrificed to collect blood for the determination of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) serum levels. Moreover, the liver was excised, weighed, and liver indices were calculated. Histopathological examination was performed to assess liver morphology. Besides, the expression of collagen type I and III in liver was determined; the mRNA expression levels of IL-6 and TNF-α in liver tissues were measured using Real-time PCR while ELISA and western blotting were performed on liver tissue homogenate to determine the protein expression of IL-6 and TNF-α. Results:The combination of praziquantel and hydroxyasiaticoside lowered the pathological scores of schistosomiasis-induced hepatic fibrosis, the liver indice, serum AST and ALT levels, improved liver morphology, downregulated the expression levels of hepatic type I and III collagen, inhibited the mRNA expression levels of pro-inflammatory factors (IL-6 and TNF-α) in the liver of mice relative to the praziquantel alone. Conclusion:The combination of hydroxyasiaticoside and praziquantel is a potential therapeutic option for schistosomiasis-induced hepatic fibrosis. Notably, this combination noticeably suppresses the protein and mRNA expression levels of pro-inflammatory factors (TNF-α and IL-6) in the liver.

Puerarin prevents vascular endothelial injury through suppression of NF-κB activation in LPS-challenged human umbilical vein endothelial cells.

OBJECTIVE: In the present study, we aimed to explore the effects of puerarin on vascular endothelial cell injury induced by lipopolysaccharide (LPS) and its underlying mechanisms. METHODS: The cell viability and morphological changes were assessed using the cell counting kit-8 (CCK-8) assay and 4´,6-diamidino-2-phenylindole (DAPI) staining, respectively. The levels of tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-1ß), monocyte/macrophage chemotactic protein-1 (MCP-1), IL-8, intercellular cell adhesion molecule-1 (ICAM-1), thrombomodulin (TM) and plasminogen activator inhibitor-1 (PAI-1) in cell culture supernatant were determined by the enzyme-linked immunosorbent assay (ELISA). The neutrophils adhesion to endothelial cells were examined by myeloperoxidase activity assay. The nuclear translocation of nuclear factor-κB p65 (NF-κB p65) was assessed by immunofluorescence analysis. RESULTS: Compared with the control group, LPS challenge significantly injured human umbilical vein endothelial cells (HUVECs) and increased the levels of TNF-α, IL-1ß, MCP-1, IL-8, ICAM-1, TM and PAI-1 in the cell culture supernatants. The neutrophils adhesion to endothelial cells were significantly increased in LPS-challenged HUVECs. Moreover, LPS challenge increased the nuclear translocation of NF-κB p65. However, puerarin pre-treatment attenuated the vascular endothelial injury and reduced the levels of TNF-α, IL-1ß, MCP-1, IL-8, ICAM-1, TM and PAI-1 in cell supernatants of LPS-challenged HUVECs. In addition, the neutrophils adhesion to HUVECs induced by LPS were also decreased by puerarin pre-treatment. Furthermore, puerarin pre-treatment reduced the nuclear translocation of NF-κB p65 elicited by LPS. CONCLUSIONS: Puerarin prevented LPS-induced vascular endothelial injury, the mechanism of which might be related to the suppression of NF-κB activation and subsequently altered levels of inflammatory factors and coagulation-related factors.

[Primary culture of human normal epithelial cells].

The traditional primary culture methods of human normal epithelial cells have disadvantages of low activity of cultured cells, the low cultivated rate and complicated operation. To solve these problems, researchers made many studies on culture process of human normal primary epithelial cell. In this paper, we mainly introduce some methods used in separation and purification of human normal epithelial cells, such as tissue separation method, enzyme digestion separation method, mechanical brushing method, red blood cell lysis method, percoll layered medium density gradient separation method. We also review some methods used in the culture and subculture, including serum-free medium combined with low mass fraction serum culture method, mouse tail collagen coating method, and glass culture bottle combined with plastic culture dish culture method. The biological characteristics of human normal epithelial cells, the methods of immunocytochemical staining, trypan blue exclusion are described. Moreover, the factors affecting the aseptic operation, the conditions of the extracellular environment, the conditions of the extracellular environment during culture, the number of differential adhesion, and the selection and dosage of additives are summarized.

Angiopoietin-like protein 3 and adiponectin levels in patients with metabolic syndrome.

OBJECTIVE: To investigate the changes of serum angiopoietin-like protein 3 (Angptl3) and adiponectin levels in patients with metabolic syndrome (MS) in order to understand their association with the MS. METHODS: Serum Angptl3 and adiponectin levels were measured by sandwich ELISA in a group of 111 patients with MS and 152 normal controls. RESULTS: Serum adiponectin was lower in the MS patients than in the control subjects [4.22(1.01-23.29) microg/mL vs. 5.41(0.97-22.27) microg/mL, P<0.05]. With regard to serum Angptl3, there was no significant difference between the 2 groups(P>0.05). Serum adiponectin was correlated to Angptl3 and high density lipoprotein-cholesterol (HDL-C)(P<0.001) and negatively correlated to body mass index (BMI), waist circumference (WC), waist to hip ratio (WHR), triglyceride (TG), fasting plasma glucose (FBG), fasting insulin (FINS) and homeostatic model assessment method-insulin resistance (HOMA-IR) (P<0.001). Serum Angptl3 was positively correlated with adiponectin (P<0.001). Serum adiponectin was found to be independently positive determinant for Angptl3 concentrations (b'=0.256, P<0.001). Adiponectin was inversely correlated with TG and HOMA-IR (b'=-0.234, -0.145, P<0.001). CONCLUSION: Adiponectin is decreased in MS patients and may be correlated to Angptl3.

Gene and protein kinase expression profiling of reactive oxygen species-associated lipotoxicity in the pancreatic beta-cell line MIN6.

Oligonucleotide microarrays were used to define oleic acid (OA)-regulated gene expression and proteomic technology to screen protein kinases in MIN6 insulinoma cells. The effects of oxidative stress caused by OA and potential protective effects of N-acetyl-L-cysteine (NAC), a scavenger of reactive oxygen species (ROS), on global gene expression and beta-cell function were investigated. Long-term exposure of MIN6 cells to OA led to a threefold increase in basal insulin secretion, a 50% decrease in insulin content, an inhibition of glucose-stimulated insulin secretion (GSIS), and a twofold increase in the level of ROS. The addition of NAC normalized both the OA-induced insulin content and ROS elevation, but it failed to restore GSIS. Microarray studies and subsequent quantitative PCR analysis showed that OA consistently regulated the expression of 45 genes involved in metabolism, cell growth, signal transduction, transcription, and protein processing. The addition of NAC largely normalized the expression of the OA-regulated genes involved in cell growth and differentiation but not other functions. A protein kinase screen showed that OA regulated the expression and/or phosphorylation levels of kinases involved in stress-response mitogen-activated protein kinase, phosphatidylinositol 3-kinase, and cell cycle control pathways. Importantly, these findings indicate that chronic OA exposure can impair beta-cell function through ROS-dependent and -independent mechanisms.

[Analysis on mutations of GJB2 gene in Chinese population].

OBJECTIVE: To determine the prevalence and types of GJB2 mutations and to investigate the genetic mechanism in Chinese autosomal recessive deafness. METHODS: The subjects were four Chinese pedigrees (39 individuals) and 50 normal adults. GJB2 was amplified by PCR. The products were digested with restriction enzyme Apa I, then sequenced. RESULTS: Homozygous deletion C at position 232-235 of GJB2 (235delC),which resulted in frameshift mutation, was found in four affected individuals of two pedigrees; the compound heterozygous deletions (235delC/232G to A) were found in two affected individuals in one pedigree. One carrier with 235delC was found in normal controls (1% allele). Two kinds of polymorphisms 79G to A(V27I) and 3 41A to G(E114G) were found in both affected and normal controls. The frequencies of allele for 79G to A and 341A to G in normal controls were 30%, 21%, respectively. CONCLUSION: 235delC mutation of GJB2 was related with Chinese autosomal recessive deafness, and the 232G to A(Ala78Thr) missense mutation was found to be a novel mutation.